A better understanding of how memory works is emerging from a
newfound ability to link a learning experience in a mouse to consequent
changes in the inner workings of its neurons. Researchers, supported
in part by the National Institutes of Health's National Institute
of Mental Health (NIMH), have developed a way to pinpoint the specific
cellular components that sustain a specific memory in genetically-engineered
mice.

"Remarkably, this research demonstrates a way to untangle
precisely which cells and connections are activated by a particular
memory," said NIMH Director Thomas Insel, M.D. "We are
actually learning the molecular basis of learning and memory."

For a memory to last long-term, the neural connections holding
it need to be strengthened by incorporating new proteins triggered
by the learning. Yet, it's been a mystery how these new proteins — born
deep inside a neuron — end up becoming part of the specific
connections in far-off neuronal extensions that encode that memory.

By tracing the destinations of such migrating proteins, the researchers
located the neural connections, called synapses, holding a specific
fear memory. In the process, they discovered these synapses are
distinguished by telltale molecular tags that enable them to capture
the memory-sustaining proteins.

Mark Mayford, Ph.D., and Naoki Matsuo, Ph.D., of the Scripps Research
Institute, report on their findings in the February 22, 2008 issue
of the journal Science.

The Scripps researchers have been applying their new technique
in a series of studies that focus on progressively finer details
of the molecular machinery of memory.

"Inside neurons involved in a specific memory, we're tracing
molecules activated by that learning to see how it ultimately changes
neural connections," explained Mayford.

In a study published in the August 31, 2007 Science,
Mayford and colleagues showed the same neurons activated by a learning
experience are also activated when that memory is retrieved. The
more neurons involved in the learning, the stronger the memory.

The researchers determined this by genetically engineering a strain
of mice with traceable neurons in the brain's fear center, called
the amygdala. Inserted genes caused activated neurons to glow red
when the animals learned to fear situations where they received
shocks, in a process known as fear conditioning — and to
glow green when the memory was later retrieved. The researchers
then chemically prevented further expression of those neurons,
so that resulting neural and behavioral changes could be confidently
attributed to that learning experience at a later time. The study
revealed which circuits and neurons were involved in the specific
learning experience.

In the new study, Mayford and Matsuo adapted this approach to
discover how fear learning works at a deeper level — inside
neurons of the brain's memory hub, called the hippocampus.

Evidence suggested that proteins called AMPA receptors (http://www.nimh.nih.gov/science-news/2007/faster-acting-antidepressants-closer-to-becoming-a-reality.shtml)
strengthen memories by becoming part of the synapses encoding them. To
identify these synapses, the researchers genetically engineered
a strain of mice to express AMPA receptors traceable by a green
glow. After fear conditioning had triggered new AMPA receptors
deep in the neuron's nucleus, they chemically suppressed any further
expression of the proteins. This allowed time for the receptors
to migrate to their appointed synapses. Hours later, green fluorescence
revealed the fate of the specific AMPA receptors born in response
to the learning.

As expected, the newly synthesized AMPA receptors had traveled
and become part of only certain hippocampus synapses — presumably
the ones holding the memory. Synaptic connections are made
onto small nubs on the neuron called spines. These spines
come in three different shapes called thin, stubby and mushroom. While
little was known about the function of these differently shaped
spines, the fact that they are altered in various forms of mental
retardation, like Fragile-X syndrome, suggests a critical importance
in mental function.

The researchers discovered the synapses that received the AMPA
receptors with memory were limited to the mushroom type. The
mushroom spines also figured prominently in the same neurons when
the fear conditioning was reversed by repeatedly exposing the animals
to the feared situation without getting shocked — a procedure
called extinction learning. This indicated that the same neurons
activated when a fear is learned are also activated when it is
lost. The surge in mushroom spine capture of the receptors appeared
within hours of learning and was gone after a few days, but appeared
to be critical for cementing the memory.

Newly synthesized AMPA glutamate receptors
(green) were captured by mushroom-shaped spines in mouse hippocampus
neurons encoding memory, after one (left), two (middle) and
six (right) hours of fear conditioning. The receptors are thought
to be key to strengthening the memory, making it long-term.
The same green fluorescent protein that makes fireflies glow
was genetically inserted into the receptors to reveal their
destinations in the brain.

The National Institute of Mental Health (NIMH) mission is to reduce
the burden of mental and behavioral disorders through research
on mind, brain, and behavior. More information is available at
the NIMH website, http://www.nimh.nih.gov.

The National Institutes of Health (NIH) — The Nation's
Medical Research Agency — includes 27 Institutes and
Centers and is a component of the U.S. Department of Health and
Human Services. It is the primary federal agency for conducting
and supporting basic, clinical and translational medical research,
and it investigates the causes, treatments, and cures for both
common and rare diseases. For more information about NIH and
its programs, visit www.nih.gov.